In the previous period of funding we have demonstrated that 1) rhabdomycosarcomas (RMS) express the myogenic regulatory factor MyoD. However, in the context of cells transformed by the expression of PAX/ALV, the function of MyoD is attenuated. Thus, MyoD expression fails to transactivate reporter genes, and fails to arrest cells in G1 phase when overexpressed. 2) We have shown also that several RMS cell lines are dependent on insulin like growth factors (Gfs) for proliferation. Inhibition of the type I IGF receptor (IGF-IR) inhibits growth, increasing the proportion of cells in G1 phase. 3) IGF-IR-dependent growth is inhibited by expression of a dominant negative p21Ras, but receptor dependence is not abrogated by oncogenic K-RAS expression. 4) Constitutive expression of IGF-IR antisense reduces tumorigenicity of RMS cells, and leads to accumulation of a high proportion of multinucleate myotube-like cells. 5) Growth of IGF-IR-dependent cell lines is highly sensitive to the inhibitory activity of the macrolide antibiotic rapamycin which induces G1 arrest and differentiation. Based upon these findings we propose to test the following hypotheses: 1. That IGF-I receptor mediated mitogenesis in RMS cells requires both transduction of signal through p21Ras, and through a distinct yet to be defined pathway. 2. That IGF-IR signaling requires mammalian TOR (Target Of Rapamycin) participation, and that mTOR represents a potential therapeutic target for therapy of rhabdomyosarcomas. 3. That IGF-IR signaling activates a kinase responsible for phosphorylation of Thr115 in the DNA binding domain of MyoD resulting in attenuated function in RMS cells. 4. That signaling through IGF-IR protects cells from cytotoxic agents, and that rapamycin may reverse this protection. We consider that the proposed studies will allow insight into new approaches to treatment of RMS, and specifically determine the potenital for rapamycin analogues for advancement to clinical trials. Studies will determine whether IGF-I receptor signaling protects RMS cells from cytotoxic agents, and the impact of blocking this on chemosensitization.